Ultrafast Redistribution of E. Coli SSB Along Long Single-Stranded DNA via Intersegment Transfer

J Mol Biol. 2014 Jun 26;426(13):2413-21. doi: 10.1016/j.jmb.2014.04.023. Epub 2014 May 2.

Abstract

Single-stranded DNA binding proteins (SSBs) selectively bind single-stranded DNA (ssDNA) and facilitate recruitment of additional proteins and enzymes to their sites of action on DNA. SSB can also locally diffuse on ssDNA, which allows it to quickly reposition itself while remaining bound to ssDNA. In this work, we used a hybrid instrument that combines single-molecule fluorescence and force spectroscopy to directly visualize the movement of Escherichia coli SSB on long polymeric ssDNA. Long ssDNA was synthesized without secondary structure that can hinder quantitative analysis of SSB movement. The apparent diffusion coefficient of E. coli SSB thus determined ranged from 70,000 to 170,000nt(2)/s, which is at least 600 times higher than that determined from SSB diffusion on short ssDNA oligomers, and is within the range of values reported for protein diffusion on double-stranded DNA. Our work suggests that SSB can also migrate via a long-range intersegment transfer on long ssDNA. The force dependence of SSB movement on ssDNA further supports this interpretation.

Keywords: optical tweezer; single-stranded DNA; single-stranded DNA binding protein.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Binding Sites
  • DNA, Bacterial / chemistry
  • DNA, Bacterial / metabolism*
  • DNA, Single-Stranded / chemistry
  • DNA, Single-Stranded / metabolism*
  • DNA-Binding Proteins / chemistry
  • DNA-Binding Proteins / metabolism*
  • Diffusion
  • Escherichia coli / metabolism*
  • Escherichia coli Proteins / chemistry
  • Escherichia coli Proteins / metabolism*
  • Microscopy, Fluorescence
  • Models, Molecular
  • Nucleic Acid Conformation
  • Optical Tweezers
  • Protein Binding
  • Protein Structure, Quaternary
  • Protein Subunits

Substances

  • DNA, Bacterial
  • DNA, Single-Stranded
  • DNA-Binding Proteins
  • Escherichia coli Proteins
  • Protein Subunits
  • SSB protein, E coli